The overarching goal of the research described in this proposal is to characterize the degree of integration of neuroendocrine mechanisms of behavior at multiple levels of biological organization, from gene/protein to hormone to behavior, among individuals and between sexes. The proposed research will probe co-variation among these levels of organization, with a focus on aggression and sex steroid hormones. Previous research provides a firm understanding of steroid action, but leaves significant uncertainty about how these levels of organization interact with one another within an individual, leading to a specific behavioral outcome. In particular, surprisingly little is known as to whether the expression of steroid hormone-binding molecules in the brain predicts variation in behavior at the level of individuals. Also not clear is whether the mechanisms that lead to individual variability in behavior within a sex are shared between the sexes, i.e. the degree to which males and females use similar or different mechanisms to produce outwardly similar behaviors. The proposed study species is a songbird, the dark-eyed junco (Junco hyemalis), which is ideal owing to extensive existing knowledge of the effects of testosterone (T) on behavioral variation among individuals and between sexes. The research will employ both experimental and observational approaches by quantifying both protein and mRNA for steroid-sensitive neural targets (androgen receptor, AR;estrogen receptor, ER;and the enzyme aromatase, ARO, which converts T to estradiol, E2) across a diversity of behaviorally relevant neural loci. Collectively, a neural systems approach is used to address two specific aims: (1) To identify patterns of co-variation between an individual's degree of aggressiveness, its level of circulating T and E2 and its neural expression of AR, ER and ARO. This aim is achieved by assaying aggression in free-living subjects and comparing their behavior to circulating hormone levels and to mRNA for neural steroid targets, using quantitative PCR. (2) To compare whether males and females use similar steroid-mediated mechanisms of aggression. This aim is achieved by using data from the first study to contrast target-behavior relationships between males and females. A second study dives deeper, using immunocytochemistry to identify sex differences in the specific neural circuitry in which one steroid target (ARO) mediates aggression. ARO-mediated aggression will be quantified by co- localizing ARO with the immediate early gene product Fos following an aggression trial. The proposed research will address a significant unknown in behavioral neuroendocrinology (individual variation), and then extend this novel approach to sex comparisons. By filling these key knowledge gaps, this research will illuminate fundamental neuroendocrine mechanisms of behavior. It will thus contribute to a foundation of basic knowledge needed for future research on neuroendocrine factors affecting steroid-mediated health issues, including sex and individual differences in the propensity for behavioral disorders with high levels of violence/ aggression (e.g. ADHD, conduct disorder, etc.), or improved treatments of related neural deficiencies. PUBLIC HEALTH RELEVANCE: By identifying the genetic and cellular factors that underlie variability in overt aggression at the individual level and comparing these neuroendocrine mechanisms between males and females, this research fills unresolved knowledge gaps about basic biobehavioral mechanisms. These studies thus lay the groundwork for future advances in the individualized treatment and prevention of a number of steroid-mediated behavioral disorders affecting children and adults of all ages. Likewise, the identification of factors that differentially promote aggression in males and females will increase understanding of fundamental sex differences in neurobiology and behavior, and could be applied to future work on the causes/treatment of known sex differences in aggressive tendencies or other hormone-mediated health issues in men and women.